BALB/c mice have previously been found to express a unique 1.7 kbp species of MMTV RNA in the lactating mammary gland. In previous studies we have established the chromosomal locations of the endogenous MMTV genomes (designated MTV-6, MTV-8, and MTV-9). Using the feral Czech II mouse strain which lacks endogenous MMTV genomes we have genetically derived sublines which contain only MTV-6, MTV-8, or MTV-9. We have found that only the MTV-6 subline expresses the 1.7 kb MMTV RNA in lactating mammary tissue. RNA:RNA in situ hybridization of sections from lactating MTG mouse mammary gland show that this viral genome is expressed in discrete regions of the gland. We have previously identified a new common integration region (designated int-3) for MMTV in MMTV (Czech II) induced mammary tumors. Recombinant clones of this region of the cellular genome have been obtained. Our recent studies have produced the following observations: (1) 6 out of 32 tumors contain a integrated viral genome at int-3, (2) in each case the transcriptional orientation of the viral genome is the same direction, (3) 5' to the integrated viral genome, cellular sequences are transcribed into a 2.4 kb species of RNA, (4) the int-3 RNA is found only in tumors with an integrated MMTV genome at this locus and is not found in normal lac mammary tissue, (5) int-3 is located 11cM from the H-2 locus on chromosome 17, with the gene order certromere-T-H-2-int-3.